The number of patients of pyorrhea (periodontal disease) and osteoporosis increase with aging. Loss of teeth or malfunction of an artificial tooth occurs by loss of the jawbone (alveolar bone) and, when the diseases advance, any implant treatments become impossible. In such situation, surgery for recovering the jawbone is conducted. Although the most effective is transplantation of autologous bone in a site of bone loss, invasion for bone collection is high risk for elderly persons and sick persons. As an alternative material, an artificial bone such as hydroxyapatite is provided. However, such artificial bone has the problems common to the inorganic materials and has a drawback in terms of formability. Although many artificial bones are currently provided in the form of granules; when the morphology of the bone defect part have no complete recess form, the risk of outflow of the granules increases. In particular for elderly persons and sick persons having lowered resistance, the outflow of the artificial bones having high fluidity with blood or external force has a risk of an important influence to the treatment process. Thus, the present inventors developed a DNA/protamine complex as a bone-repairing material excellent in workability, which is difficult to flow and yet capable of being directly injected through a syringe and easily moldable into a free shape (Patent Literature 1).
A protamine as a basic protein obtained from the testes (milt) of fish is reported to have an activity of inhibiting Arg-gingipain as a protease produced by Porphyromonas gingivalis (see Non Patent Literature 1). In addition, it is disclosed that periodontitis caused by a periodontal pathogen can be suppressed by further adding a protamine or a derivative thereof, and tranexamic acid and/or epsilon aminocaproic acid (see Patent Literature 2). These protamines are commonly used as a food preservative and are highly safe; thus, protamine is a promising medicine as an alternative to the conventional antimicrobial agents and antibiotics. Further, it is disclosed that protamine has a suppressing effect on growth of Streptococcus mutans, which is a caries-causing bacterium (see Patent Literature 3) and a suppressing effect on the adhesion of oral bacteria (see Patent Literature 4).
The present inventors have found that a protamine hydrolysate exhibited an antifungal activity against fungi, such as the genus Candida, causing oral candidosis (see Patent Literature 5). In addition, the present inventors have found that a protamine also had an anti-bacterial activity against periodontal disease bacteria, such as Porphyromonas gingivalis and Prevotella intermedia (see Patent Literature 1).
Protamine is a kind of polycations, and it is known that it forms a water-insoluble complex by generating an electrostatic binding with an anionic high molecular compound.
DNA obtained from the fish testes in the same manner as protamine has many phosphate groups providing anionic binding factors in the DNA molecule. Since such DNA exhibits an electrostatic affinity to a cationic substance; the DNA can form an electrostatic reactant with the cationic substance. It is disclosed that, although DNA molecule is poor in formability in itself and has a difficulty in control of its diffusion velocity through in vivo metabolism due to its water-solubility, a water-insoluble and self-supporting transparent film can be prepared by electrostatically binding a DNA molecule to an artificial cationic lipid (see Patent Literature 6).
A film consisting of a complex of DNA and a lipid is also known. The DNA/lipid complex film, in which the DNA molecule as a biopolymer keeps a regular double helix structure, enables various low-molecular compounds to intercalate into the spaces between DNA bases in the double helix structure, and similar low-molecular compounds to groove-bind in the two grooves (main and minor grooves) of the DNA. A method of producing a medical material by intercalating and/or groove-binding a medicinal ingredient to the DNA/lipid complex film was invented (see Patent Literature 7).
The present inventors focused attention on the fact that chitosan already used as a medical and/or dental material was a cationic substance and, thus, the present inventors made an invention based on that a complex of a DNA molecule and chitosan held a double helix structure specific to DNA, insolubility in water, biocompatibility, antibacterial activity and a good formability (see Patent Literature 8). The present inventors also invented a method for easily molding a DNA/chitosan complex into a thread form, a ball form, or a disk form by suspending it in a phosphate buffer solution (see Patent Literature 9); and a method for producing a film by compression-molding such a complex under heating (see Patent Literature 10). The DNA/chitosan complex and its molded products have been a promising one as a medical and/or dental material, because it had an excellent biocompatibility and biostability.
There are the prior literatures relate to elastin, which is present in the elastic tissues of the body, such as the ligament, blood vessels, the heart, and the skin, and which activates transforming growth factor-β1 (TGF-β1) and induces bone formation reaction (see Non Patent Literatures 2 and 3). There are two type of elastin from Elastin Products Company, Inc., which were used in these prior literatures, i. e., α-elastin having a molecular weight of 6,600 to 47,000 Da, extracted from the nature, and an elastin peptide having a molecular weight of 1,000 to 25,000 Da, obtained by degradation by acid and/or alkali treatment. Alpha-Elastin from Elastin Products Company, Inc. can be obtained by dissolving a bovine neck ligament by a high temperature oxalic acid, separating and removing β-elastin by a coacervation method after purification and desalting. The acid- and/or alkali-treated elastin can be obtained by washing a bovine neck ligament with hot water to remove soluble contaminating proteins, boiling the resultant in an alkaline solution, and hydrolyzing and purifying the insoluble fraction. Both of these elastins are water soluble.
However, although the effect of elastin to induce bone formation by the in vitro investigation has been recognized, it is necessary that elastin has formability or that elastin is included in another material. Even if elastin is contained in a prior art material by impregnation, dissolution and outflow of elastin in vivo in a very short time has been expected. Therefore, elastin has not been considered to exhibit sufficient bone formation ability. The elastin can be cross-linked using a crosslinking agent, etc., to provide formability to elastin. However, it is desirable to minimize the use of an agent such as a crosslinking agent in a bone formation material because there is concern that the crosslinking agent itself causes a reaction against a foreign material or a toxic reaction in vivo.